CEACAM6 (MSVA-066R)

CEACAM6 (CD66c) is a glycosylphosphatidylinositol (GPI)‑anchored cell‑surface protein of the carcinoembryonic antigen (CEA) immunoglobulin superfamily. The CEACAM6 gene maps to chromosome 19q13.2 within the CEA gene cluster. CEACAM6 mediates cell–cell adhesion (homo‑ and heterophilic interactions with other CEACAMs), and contributes to epithelial architecture and innate immune interactions. It is primarily expressed in granulocytes, alveolar macrophages and in selected epithelial cell types. Genetic diseases or cancer predispositions are not known to be related to mutations or variants of the CEACAM6 gene. Unlike other cell adhesion molecules, CEACAM6 lacks both a transmembrane and a cytoplasmic domain. This enables CEACAM6 to mediate cell-cell and cell-matrix adhesion through lateral associations with lipid rafts and co-receptors, especially integrins. Evidence for a role of CEACAM6 in cancer is accumulating. CEACAM6 is commonly upregulated in epithelial malignancies and high expression levels have been found to be associated with invasion, metastasis, and chemotherapy resistance. Based on its location on the cell‑surface CEACAM6 is currently being explored as a therapeutic target protein in cancer.

CEACAM6 staining predominates in subsets of inflammatory cells including granulocytes and alveolar macrophages. Significant CEACAM6 expression also occurs in epithelial cell types such as for example pneumocytes of the lung. Images describing the CEACAM6 staining pattern in normal tissues obtained by the antibody MSVA-066R are shown in our “Normal Tissue Gallery”.

These findings are largely comparable to the RNA and protein data described in the Human Protein Atlas (Tissue expression CEACAM6).

Positive control = Lung: A strong membranous CEACAM6 staining of all alveolar pneumocytes and a strong cytoplasmic and membranous CEACAM6 positivity of alveolar macrophages should be seen.

Negative control = Kidney: All epithelial cells must not show CEACAM6 staining.

Normal tissue gallery

CEACAM6 expression is especially common in pancreatic adenocarcinoma and in other gastrointestinal epithelial cancers. It can occur in other cancer types.

The TCGA findings on CEACAM RNA expression in different tumor categories have been summarized in the Human Protein Atlas.

Cancer tissue gallery

IHC users have different preferences on how the stains should look like. Some prefer high staining intensity of the target stain and even accept some background. Others favor absolute specificity and lighter target stains. Factors that invariably lead to more intense staining include higher concentration of the antibody and visualization tools, longer incubation time, higher temperature during incubation, higher temperature and longer duration of the heat induced epitope retrieval (slide pretreatment). The impact of the pH during slide pretreatment has variable effects and depends on the antibody and the target protein. Accordingly, multiple different protocols can generate identical staining results.

All images and data shown here and in our image galleries are obtained by the manual protocol described below. Other protocols resulting in equivalent staining are described as well.

Manual protocol

Freshly cut sections should be used (less than 10 days between cutting and staining). Heat-induced antigen retrieval for 5 minutes in an autoclave at 121°C in pH9 Target Retrieval Solution buffer. Apply MSVA-066R at a dilution of 1:150 at 37°C for 60 minutes. Visualization of bound antibody by the EnVision Kit (Dako, Agilent) according to the manufacturer’s directions.

• Knowledge on prevalence and clinical significance of CEACAM6 expression across different cancer types is incomplete.
• Are there tumor specific structural/glycosylation variants of CEACAM6 that can be exploited for highly tumor‑selective therapeutics? 
• How does CEACAM6 expression modulate anti‑tumor immunity in human tumors? 
• Can CEACAM6 be used as an early detection marker for pancreatic neoplasms? 
• Is tumor‑cell CEACAM6 expression spatially relate with local immune exclusion?

There are two ways how the specificity of antibodies can be documented for immunohistochemistry on formalin fixed tissues. These are: 1. Comparison with a second independent method for target expression measurement across a large number of different tissue types (orthogonal strategy), and 2. Comparison with one or several independent antibodies for the same target and showing that all positive staining results are also seen with other antibodies for the same target (independent antibody strategy). 

Orthogonal validation: For the antibody MSVA-066R specificity is supported by the result of a comparison of its staining pattern on 76 normal tissue types with CEACAM6 RNA expression data in normal tissues which were collected in three independent RNA screening studies, including the Human Protein Atlas (HPA) RNA-seq tissue dataset, the FANTOM5 project, and the Genotype-Tissue Expression (GTEx) project, and which are summarized in the Human Protein Atlas (Tissue expression CEACAM6).  Consistent with RNA expression data, CEACAM6 immunostaining by MSVA-066R was seen in all tissues for which RNA expression had been described including lung, gastrointestinal tract, gallbladder, cervix uteri, bone marrow, appendix, and the tonsil. Also in line with RNA data, immunostaining was regularly absent in tissues for which CEACAM6 RNA expression had not been described such as the kidney, prostate, testis, epididymis, seminal vesicles, brain, endocrine organs, endometrium, ovary and the placenta.

Comparison of antibodies: True expression of CEACAM6 in all cells with CEACAM6 positivity by MSVA-066R is further corroborated by a confirmation of each individual staining seen by MSVA-066R in 76 different normal tissue types by a commercially available independent second antibody (termed “validation antibody”). An additional cytoplasmic staining of epithelial cells in the gallbladder and the uterine endocervix which was only seen by the validation antibody and not by MSVA-066R was considered an antibody specific cross-reactivity of the validation antibody.

Normal tissue gallery